This report identifies L-ethionine as an inducer of differentiation in murine erythroleukemia cells. When Friend erythroleukemia cells are grown in the presence of 4 mM L-ethionine, globin mRNA accumulates and in 4-5 days, 25 -30 of the cells in the culture contain hemoglobin. Incubation of the cells with bromodeoxyuridine prevents both ethionine-induced accumulation of globin mRNA and er y thr oid differentiation.At the concentration where L-ethionine acts as an inducer of FL cell differentiation it inhibits methylation of DNA and tRNA in vivo but does not prevent macromolecular synthesis or cell division.To establish whether a link existed between inhibition of a specific methyltransferase and activation of globin synthesis in FL cells, we examined the degree of hypomethylation of DNA and tRNA from F L cells induced to differentiate with dimethylsulfoxide and butyrate.In contrast to the tRNA from ethionine-treated cells, tRNA from cells induced by butyrate or MezSO cannot be methylated in vitro using homologous enzymes.DNA isolated from cells exposed to any of the three inducers, however, was significantly hypomethylated when compared with DNA from uninduced cells. These data suggest that methylation of DNA may play a role in the regulation of gene expression.
There were no significant differences between the mean blood plasma (leu-kocyte-free) RNAuse activity among 128 healthy women volunteers age 13-70 and 49 women with benign gynecological tumors. Exceptions to this finding were three apparently healthy women volunteers who had plasma enzyme activity which was higher than two standard deviations from the mean of the control subjects. Increased plasma RNAase activity was also demonstrated for 21 of 22 patients with ovarian carcinomas of different histological types. This group included two patients with Stage IA, two patients with Stage IC ovarian carcinoma, and 17 patients with advanced ovarian carcinoma. The one exception was a patient with a well encapsulated, mucinous cystadenocarcinoma, Stage IA. The plasma RNAuse activity returned to normal values in all of the cancer patients who had no clinical evidence of residual malignant tissue after surgical treatment. However, the enzyme activity also returned to a normal value in one of the 17 women in whom all of the malignant tissue was not removed. These data indicate that plasma RNAase activity can be utilized as a tumor marker for the presence of ovarian malignancies of various histological types, and to differentiate between malignant and benign neoplasms.0 oratory is to establish a rapid and reliable method for the early diagnosis of ovarian cancer, ovarian cancer being the foremost problem in gynecological cancer we have today. In spite of the extensive use of surgery, radiotherapy and/or chemotherapy, the overall survival rate has remained approximately 30% over the past 30 years. g,21 The poor prognosis is largely attributable to the fact that at the time of diagnosis about 70% of ovarian cancers already have widespread intraperitoneal metastases. From these data it is evident that a practical method for the early diagnosis of ovarian cancer is essential. In order to achieve this goal, we elected to study the possibility of relating the presence of ovarian malignancies with the appearance of elevated blood serum or plasma ribonuclease (RNAase) activity. The rationale for this approach is based on the findings of other in-From the
The existence of a DNA-dependent protein methylase activity without any concomitant DNA methylase activity was demonstrated in bull seminal plasma. The enzyme utilized S-adenosyl-L-methionine as a methyl donor, and endogenous seminal plasma protein as the substrate. There was no demonstrable enzyme activity when the seminal plasma was preheated at 100 degrees for 10 min, or when the enzyme reaction mixture was incubated at 4 degrees. The protein methylase required a heterologous DNA source, had optimal activity at pH 8.1, and was enhanced in the presence of Mg2+, NH4+, and reduced glutathione. After the methylated protein product was separated from DNA by extraction with 0.2 M HCl, the incorporated radioactivity was shown to be totally solubilized by incubating the protein either with Pronase or 1 M NaOH, while RNase and DNase had no effect. Approximately 70% of the enzymatically synthesized amino acids in the protein product were tentatively identified as O-methylated amino acid ethers by virtue of their elution from a Dowex 50 H+ column with 0.2 M pyridine, and their stability to acid and base hydrolysis. The partially purified methylated product was shown by Sephadex G-50 chromatography to consist of three distinct radioactive proteins with molecular weights of approximately 21,000, 15,000, and 10,000.
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